DESCRIPTION (Verbatim from Applicant's Abstract): The long-term objective of this research is to determine the multiple roles of fluid wall shear stress (blood flow) on the water and solute transport barrier properties of the endothelial layer which lines all blood vessels. Shear-dependent endothelial transport has important implications for the normal function of microvessels which must deliver material to tissue in proportion to blood flow. In this context, an increase in shear stress is expected to increase transport. On the other hand, shear-dependent endothelial permeability to macromolecules such as low density lipoprotein has been hypothesized to play a key role in the localization of atherosclerotic lesions in regions of low and oscillatory shear in arteries. Regions of unidirectional high shear tend to be spared. In this scenario, an increase in shear is hypothesized to reduce transport. In the proposed research, a unique shearing apparatus will be used to explore the mechanisms controlling the shear stress response of transport properties in two in vitro cell culture models: bovine aortic endothelial cells (BAE(s)) which display an increase in transport in response to shear stress, and human umbilical vein endothelial cells (HUVECs) which show the opposite response. The specific aims of the proposed research are: 1. To determine the effects of steady and oscillatorv shear stress on the hydraulic conductivitv (Lp) and macromolecular permeabilitv (Pe) of HUVEC and BAEC monolavers. We will culture HUVEC and BAEC monoiayers on porous filters and measure Lp and Pe in response to steady shear stress and oscillatory shear stress (with orwithout reversal) in a unique apparatus developed in our laboratory which allows us to assess Lp and Pe (multiple solutes) simultaneously. Solutes to be studied include albumin, LDL and 2000 kD dextran in experiments of 4 hours duration. 2. To determine the physical transport pathways that are affected bY shear stress. The basic hypotheses to be tested are that shear stress upregulates the formation of vesicles which transport large solutes across the endothelium, and that shear stress alters tight junction proteins in the interceilular junction pathway which accommodates water transport. 3. To determine the biochemical mechanisms mediatina the effects of shear stress on EC transport. The basic hypotheses to be tested are that BAECs and HUVECs follow different biochemical signaling mechanisms to altertransport properties in response to shear stress, and that for each cell type intercellular junctions and vesicles are controlled by different signaling mechanisms in response to shear stress. We will conduct experiments using activators and inhibitors of b aboutochemical pathways within cells to probe for the mechanisms underlying the shear stress responses of transport properties in BAECs and HUVECs.